Explain why a restriction endonuclease and DNA ligase are both essential when inserting a human gene into a bacterial plasmid to produce a recombinant plasmid.
Written & reviewed by James Millett — Biology (Imperial College London), PGCE Science (University of Cambridge).
Genetic engineers routinely use bacterial plasmids as vectors to introduce human genes into bacterial cells. This allows bacteria to produce human proteins such as insulin on an industrial scale.
Model answer (4 marks)
A restriction endonuclease recognises a specific sequence on the plasmid and cuts the DNA, producing complementary sticky ends. The same enzyme cuts the donor human gene at the same recognition sites, giving the gene fragment identical sticky ends. These complementary sticky ends allow the human gene to anneal to the plasmid by hydrogen bonding.
DNA ligase then seals the nicks in the sugar‑phosphate backbone, forming covalent phosphodiester bonds and permanently joining the gene into the plasmid.
DNA ligase then seals the nicks in the sugar‑phosphate backbone, forming covalent phosphodiester bonds and permanently joining the gene into the plasmid.
Examiner tips
- Use the word "restriction endonuclease" and "DNA ligase" explicitly
- Explain that the enzyme cuts to give sticky ends and that ligase seals the backbone
- Show the sequence of events: cut → anneal → ligate
Common mistakes
- Confusing restriction endonuclease with DNA polymerase
- Saying the enzyme only cuts the plasmid but not the donor DNA
- Forgetting to mention that ligase forms phosphodiester bonds
Mark scheme (4 marks)
- Restriction endonuclease cuts the plasmid (and the donor DNA) at specific recognition sequences
- This produces complementary sticky ends on both the plasmid and the human gene fragment
- The complementary sticky ends allow the human gene to anneal/hydrogen bond into the plasmid
- DNA ligase seals the sugar-phosphate backbone by forming covalent/phosphodiester bonds, permanently joining the gene into the plasmid
Key terms in this question
restriction endonuclease · plasmid · recombinant plasmid · DNA ligase
Related
- All OCR A-Level Biology A (H420) revision notes →
- How to answer a "Explain" question →
- Decode the mark scheme abbreviations →
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